Method for removing horn substances from animal skin

ABSTRACT

The invention relates to a method for removing horn substances from animal skin. The method is characterized in that animal skins are treated in an aqueous bath containing between 0.05 and 5 wt. %, in relation to the weight of the salt, of at least one compound of general formula I or the corresponding alkali metallic salts or alkaline-earth metallic salts thereof or ammonium or phosphonium salts. The aqueous bath also comprises at least one compound which catalyses the hydrolysis of peptide compounds

The present invention relates to a process for removing horny substancesfrom animal hides, wherein animal hides are treated in an aqueous liquorcomprising from 0.05 to 5% by weight, based on the salted weight, of oneor more compounds of the formula I

or the corresponding alkali metal or alkaline earth metal salts orammonium or phosphonium salts thereof, the variables being defined asfollows:

-   R¹ is selected from hydrogen and C₁-C₁₂-alkyl, unsubstituted or    substituted by one or more S—H or O—H groups;-   X¹ to X⁴ are identical or different and are selected from hydrogen,    C₁-C₄-alkyl, O—H, S—H and N—HR²,-   R² is hydrogen or C₁-C₁₂-alkyl or a C₁-C₄-alkyl-C═O group,

at least one of the radicals X¹ to X⁴ being S—H,

and, if R¹ contains neither O—H nor S—H, at least one further radicalfrom among X¹ to X⁴ is selected from S—H, OH and NH—R²,

and furthermore comprising at least one compound which catalyzes thehydrolysis of peptide bonds.

Animal hides have been processed to leather since antiquity. Before itis possible to begin the actual leather production, the tanning, theanimal hides must be prepared. This preparation generally takes place inthe beam house and comprises numerous operations. Most of theseoperations serve for separating off those components of the animal hideswhich are undesired in the subsequent leather production or in thesubsequent leather. The undesired components generally include, forexample, the hairs together with the hair roots. The unhairing of theanimal hides is usually assisted by chemicals. A distinction is madebetween oxidative, reductive and enzymatic unhairing methods. Anoverview of methods can be found in Herfeld, Bibliothek des Leders, Vol.2, 1988, pages 62-167 and in E. Heidemann, Fundamentals of LeatherManufacturing, E. Roether K G Druckerei und Verlag, Darmstadt 1993,pages 165-218.

In general, the unhairing of the animal hides is effected substantiallyor completely during the liming or the painting. Conventional unhairingreagents which are advantageous in production are Na₂S and NaSH, thelatter often also being referred to as sodium sulfhydrate. Both saltscan be used in highly contaminated form, and technical-grade Na₂Sgenerally has an Na₂S content which does not exceed 65% by weight andtechnical-grade NaHS usually contains 70-72% by weight of NaHS. Both,Na₂S and NaHS, have disadvantages in practical use. For safety reasons,Na₂S and NaHS can be used only in a strongly alkaline medium because,under acidification, they evolve toxic and foul-smelling hydrogensulfide. For ecological and process engineering reasons, the eliminationof the unconsumed sulfide, in particular of the sulfide-containingwastewaters, is a problematic step. If excess sulfide is precipitated,for example with Fe²⁺ or Fe³⁺, iron sulfide sludges which are expensiveto separate off are obtained. It is also possible to attempt to convertsulfides into ecologically safe salts by oxidation with, for example,H₂O₂, but corrosion problems then have to be accepted.

There has therefore been no lack of attempts to use reagents other thanNa₂S or NaHS for the treatment of the animal hides. Most experimentsstart from volatile SH— containing organic reagents.

U.S. Pat. No. 1,973,130 describes the use of organic sulfur compounds inthe presence of lime, (column 1, line 40) for unhairing, for example,calf hides. In particular, ethyl mercaptan is a foul-smelling reagentand ethyl mercaptan-containing wastewaters are difficult to work up,preventing the use of ethyl mercaptan in the beam house.

FR 1.126.252 describes the unhairing of animal hides by the action ofthioglycolamide (example 1) or thioglycerol (example 2) in the presenceof ammonium sulfate at a pH of 7-8.

However, attempts to substitute Na₂S or NaHS by mercaptoacetic acid ormercaptoethanol or the alkali metal or alkaline earth metal saltsthereof did not lead to success because both reagents and also theiralkali metal and alkaline earth metal salts readily eliminate hydrogensulfide and have an extremely unpleasant smell. Furthermore, beam housewastewaters comprising mercaptoacetic acid or mercaptoethanol ordecomposition and secondary products are difficult to clarify and giveup unpleasant odors.

The use of 1,4-dimercaptobutanediol-containing formulations for removinghorny substances, in particular hairs, from living tissue is known fromthe cosmetics industry, for example for undesired beard growth. Thus, DE21 31 630 shows that compositions comprising at least 0.25% by weight ofdimercaptobutanediol and from about 0.01 to 40% by weight of awater-soluble guanidine compound and having a pH of less than 12 can beapplied to guinea pigs in order to unhair them or to human horny skin inorder to eliminate calluses, without causing skin irritations in guineapigs or even erythremia (malignant proliferations of the formativesystem of the red blood corpuscles). The epidermis is preserved in thetreatment described in DE 21 31 630.

EP-A 0 095 916 discloses the use of formulations comprisingaminoethanethiol and 1,4-dimercaptobutanediol and an aminoguanidine ordiguanidine compound for eliminating undesired human body and facialhair. On page 2, line 1, it is stated that small thiol molecules arepreferably suitable for producing rapid unhairing because they penetratemore rapidly into the skin. The epidermis is preserved in the treatmentdescribed in EP-A 0 095 916.

EP-A 0 096 521 discloses the use of formulations comprising, forexample, 1,4-dimercaptobutanediol and an aminoguanidine or diguanidinecompound for eliminating undesired human body and facial hair. Theepidermis is preserved in the treatment described in EP-A 0 096 521.

It is furthermore known that collagen can be modified by opening S—Sbridges in the collagen by reaction with dithioerythrol and subsequentchlorination with chloroacetamide or chloroacetic acid, cf. for exampleE. Heidemann, Fundamentals of Leather Manufacturing, E. Roether K GDruckerei und Verlag, Darmstadt 1993, page 253. Furthermore, proteinsolutions can be conserved by adding dithioerythrol or dithiothreitol.The conservation is based on a type of protection from oxidation,because dithioerythrol is usually the first to be oxidized instead ofthe protein SH groups.

DE 29 17 376 C2 discloses that animal hides can be unhaired usingenzymes in the presence of compounds of the formula A1 or A2

Here, R′ are selected from hydrogen, an amino group and alkyl radicalsof 1 to 6 carbon atoms, n is from 0 to 6 and R″ is an alkyl radical of 1to 6 carbon atoms. The animal hides are treated first in the acidic pHrange with thioglycolic acid (example 1), mercaptoacetic acid (example2) or mercaptoethanol and thioglycolic acid (example 3) or a combinationof thioglycolic acid and thiourea. However, the pretreatmentcompositions have a very unpleasant odor.

WO 96/19560 proposes unhairing cattle hides by means of 2 differentenzymes and dithiothreitol (example 2, page 14, lines 10 to 12), thehairs being preserved; however, no instructions for carrying out theproposed process are disclosed.

It is an object of the present invention to provide a process forremoving horny substances from animal hides and for removing theepidermis very substantially in the same operation, in which as few aspossible of unpleasant odors are given off. In particular, it is anobject of the present invention to provide a process for removing hornysubstances so that they are very substantially destroyed.

We have found that this object is achieved and that the process definedat the outset is very suitable for removing horny substances from animalhides and for removing the epidermis very substantially in the sameoperation, and that the reagents used give off few or no unpleasantodors.

In the context of the present invention, horny substances are understoodas meaning calluses, feathers, nail and claw parts and in particularhairs of animals.

The animal hides may contain residues of flesh of the relevant animals.What is essential to the invention, however, is that they contain hornysubstances. The amount of horny substance, based on the total weight ofthe animal hide, is not critical. The novel process is suitable both forremoving large amounts of horny substance and for removing small hairresidues.

In the context of the present invention, animal hides are understood asmeaning not only hides of slaughtered animals or animals deliberatelykilled in another manner but also hides of those animals which have diedas a result of accidents, for example traffic accidents or fights withmembers of the same species or other animals, or through natural causes,such as age or disease.

In the context of the present invention, the animal hides are usuallyhides of vertebrates, e.g. cattle, calves, pigs, goats, sheep, lambs,elks, game, such as stags or does, and furthermore birds, for exampleostriches, fish or reptiles, such as snakes.

The following procedure is advantageously followed for carrying out thenovel process.

Animal hides are treated with from 0.05 to 5% by weight, based on thesalted weight, of one or more compounds of the formula I

or the corresponding alkali metal or alkaline earth metal salts orammonium or phosphonium salts, the radicals in formula I being definedas follows:

-   R¹ is selected from    -   C₁-C₁₂-alkyl, such as methyl, ethyl, n-propyl, isopropyl,        n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl,        sec-pentyl, neopentyl, 1,2-dimethylpropyl, isoamyl, n-hexyl,        isohexyl, sec-hexyl or n-decyl, particularly preferably        C₁-C₄-alkyl, such as methyl, ethyl, n-propyl, isopropyl,        n-butyl, isobutyl, sec-butyl and tert-butyl;    -   C₁-C₁₂-alkyl, substituted by or more hydroxyl or thiol groups,        such as hydroxymethyl, 2-hydroxyethyl, 1,2-dihydroxyethyl,        3-hydroxy-n-propyl, 2-hydroxyisopropyl, ω-hydroxy-n-butyl,        ω-hydroxy-n-decyl, HS—CH₂—, HS—(CH₂)₂— or HS—(CH₂)₃—,    -   and very particularly preferably hydrogen,-   X¹ to X⁴ are identical or different and are selected from hydrogen,    -   C₁-C₄-alkyl, such as methyl, ethyl, n-propyl, isopropyl,        n-butyl, isobutyl, sec-butyl and tert-butyl,    -   O—H, S—H or N—HR², in particular O—H or S—H,-   R² is hydrogen or    -   C₁-C₁₂-alkyl, such as methyl, ethyl, n-propyl, isopropyl,        n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl,        sec-pentyl, neopentyl, 1,2-dimethylpropyl, isoamyl, n-hexyl,        isohexyl, sec-hexyl or n-decyl, particularly preferably        C₁-C₄-alkyl, such as methyl, ethyl, n-propyl, isopropyl,        n-butyl, isobutyl, sec-butyl and tert-butyl;    -   or H—C═O or a C₁-C₄-alkyl-C═O group, for example acetyl,        C₂H₅—C═O, n-C₃H₇—C═O, iso-C₃H₇—C═O, n-C₄H₉—C═O, iso-C₄H₉—C═O,        sec-C₄H₉-C═O, tert-C₄H₉—C═O.        in the presence of at least one compound which catalyzes the        hydrolysis of peptide bonds.

At least one of the radicals X¹ to X⁴ is S—H and, if R¹ contains neitherO—H nor S—H, at least one further radical from among X¹ to X⁴ isselected from S—H, OH and NH—R².

Preferably, at least one group X¹ to X⁴ is hydroxyl and particularlypreferably at least two groups X¹ to X⁴ are hydroxyl. Very particularlypreferably, X² and X³ are each hydroxyl. Very particularly preferably,X¹ and X⁴ are each S—H and very particularly preferably R¹ is hydrogen.

Among the corresponding alkali metal and alkaline earth metal salts, inparticular the mono- and disodium salts, mono- and dipotassium salts andpotassium sodium salts of the compounds of the formula I may bementioned, and furthermore the corresponding calcium and magnesiumsalts. The ammonium salts and primary, secondary, tertiary and inparticular quaternary mono- and diammonium salts and phosphonium saltsmay also be mentioned. Of course, mixtures of compounds of the formula Iand the corresponding alkali metal or alkaline earth metal salts orammonium or phosphonium salts thereof can also be used.

Preferred mono- and diammonium salts have, as cations, those of theformula N(R³)(R⁴)(R⁵)(R⁶)⁺, where R³ to R⁶ are in each case identical ordifferent and are selected from hydrogen C₁-C₁₂-alkyl, phenyl andCH₂—CH₂—OH. Examples are tetramethylammonium, tetraethylammonium,methyldiethanolammonium and n-butyldiethanolammonium. Preferred mono-and diphosphonium salts have, as cations, those of the formulaP(R³)(R⁴)(R⁵)(R⁶)⁺, where R³ to R⁶ are as defined above.

Very particularly preferably, one or more 1,4-dimercaptobutanediols,selected from I a, I a′ and I b,

or the corresponding alkali metal or alkaline earth metal salts thereofare used. I a and I a′ are referred to as dithiothreitol, and I b isalso referred to as dithioerythrol. The use of racemic dithiothreitol isvery particularly preferred. I a, I a′ and I b are virtually odorless,readily meterable and readily water-soluble compounds.

The compounds I a or I a′ and I b are known and are commerciallyavailable, for example, from Aldrich or ACROS Chemicals. Further membersare synthesized as described in U.S. Pat. No. 4,472,569 or J. Chem. Soc.1949, 248 or by analogous reactions.

At least one novel process is carried out in the presence of at leastone compound which catalyzes the hydrolysis of peptide bonds.

At least one of these compounds is preferably an organic compound.

In the context of the present invention, compounds which catalyze thehydrolysis of peptide bonds are not to be understood as meaning Brönstedacids or the salts thereof.

In the context of the present invention, organic compounds whichcatalyze the hydrolysis of the peptide bonds are to be understood inparticular as meaning enzymes. Exo and endopeptidases are preferred.These may be members of the main classes of proteases, for exampleserine proteases, cysteine proteases, metalloproteases and acidproteases.

An enzyme may be used.

Mixtures of 2 enzymes may be used.

Examples of serine proteases are trypsin, chymotrypsin, elastase,thrombin, plasmin, subtilisin and acrosin.

Examples of cysteine proteases are papain, bromelain and cathepsin B.Examples of metalloproteases are carboxypeptidase and ACE (angiotensinconverting enzyme).

Examples of acid proteases (aspartate proteases) are pepsin and HIVprotease.

Serine proteases, for example trypsin, chymotrypsin, subtilisin andproteinase K, and variants of the abovementioned enzyme are particularlysuitable for the purposes of the present invention. Variants include,inter alia, mutants which have formed as a result of insertion(s),deletion(s) and point mutation(s) and have changed, in particularadvantageous, properties in comparison with the protease used as astarting material in each case. Examples of changed properties arethermal stability, higher affinity to the substrate to be convertedenzymatically, (higher) substrate specificity and a shift of the optimumpH into the desired pH range. In the context of the present invention,fragments of abovementioned proteases are also referred to as variants.The variants are prepared by a recombinant method using the conventionalmethods, e.g. those described in Molecular Cloning—A Laboratory Manualby Sambrook, Fritsch and Maniatis (1989), in a suitable bacterial orfungal host system. Proteases of the four main classes (serineproteases, cysteine proteases, metalloproteases and acid proteases)having specific keratinolytic activity and mixtures of these enzymes arevery particularly preferred. In the context of the present invention,enzymes which hydrolyze peptide bonds are also to be understood asmeaning commercially available enzyme formulations. Examples of suchproducts are Alcalase 3.0T, Pyrase 250 MP, concentrated PTN 3.0 (type p)from Novozymes, Prozym 6 from TFL, pancreatin from Nordmark A,Pancreatina enzyme conc. from Scientific Protein Laboratory, Alprolase 3m, Basozym® L10 and Basozym® S20 from BASF-Aktiengesellschaft, Batinase(producer: Genencor), Proleather (producer: Amano), Protease L 660(producer: Genencor), Esperase (producer: Novo Nordisk), Alcalase 2.4L(producer: Novo Nordisk), Savinase (producer: Novo Nordisk) and Pruafect4000 L (producer: Genencor).

If the abovementioned enzymes or variants of these enzymes are usedalone or as mixtures in the novel process, not only is particularly goodremoval of horny substances achieved but also substantial or preferablycomplete degradation of the epidermis is observed.

In general, an amount of from 0.05 to 5% by weight, based on the hideweight or salted weight of the animal hides, of compound I issufficient. From 0.1 to 1.5% by weight is preferred and from 0.25 to1.0% by weight is particularly preferred.

The amount in which the compound which catalyzes the hydrolysis ofpeptide bonds is used, in particular the amount of enzyme used, isusually expressed in Löhlein-Volhard units (LVUs). Usually, instead ofmetering pure enzyme, dilute formulations which may be solid or liquidare metered.

The LVUs are determined by titrimetric methods which are known per seand are based on the degradation of casein by an enzyme formulation tobe investigated or an enzyme to be investigated and subsequent titrationof the liberated carboxyl groups with 0.1 N NaOH.

One LVU is equivalent to 0.00575 ml of 0.1 N NaOH.

According to the invention, from 500 to 2 000 000, preferably from 1 000to 50 000, particularly preferably from 1 500 to 10 000, LVU/kg, basedin each case on the salted weight of the animal hide to be treated, aremetered.

A compound or compounds which catalyzes or catalyze the hydrolysis ofpeptide bonds is or are used as a rule in amounts which are at least afactor of 10, preferably 100, particularly preferably 1 000, smallerthan the amount of compound I, based on pure compounds.

Particularly if the compound used is one or more enzymes, instead of thepure enzyme usually one or more solid or liquid formulations whichcontain a compound which catalyzes the hydrolysis of peptide bonds ismetered.

Solid formulations contain, in addition to the compound or the compoundswhich catalyzes or catalyze the hydrolysis of peptide bonds, alsoinorganic or organic solids or mixtures thereof. Examples of inorganicsolids are NaCl, Na₂SO₄, kieselguhr, NaHCO₃, Na₂CO₃ or kaolin,bentonites or clay minerals; suitable organic solids are, for example,polysaccharides, such as starch and modified starch, or urea. Solidformulations may furthermore contain reducing substances, for exampleNaHSO₃. Liquid formulations contain at least one liquid solvent ordispersant, for example water or a mixture of water and organic solvent.

The novel treatment of the animal hides is preferably effected with oneor more compounds of the formula I and at least one compound whichcatalyzes the hydrolysis of peptide bonds, during liming or duringpainting, in particular under either hair-destroying or underhair-preserving conditions. Instead of the conventional concentration offrom about 2 to 4% by weight of Na₂S or NaHS, it is possible duringliming or during painting to manage with a concentration of less than0.1% by weight of Na₂S or NaHS for an equivalent effect with regard tothe removal of horny substances. In a preferred variant of the novelprocess, it is possible to dispense with the use of Na₂S or NaHS orother foul-smelling sulfur-containing reagents.

According to the invention, the animal hides are treated in an aqueousliquor. The liquor ratio is from 1:10 to 10:1, preferably from 1:2 to4:1, particularly preferably up to 3:1, based on the hide weight orsalted weight of the animal hides.

The novel process is carried out at a pH of from 6 to 14, preferablyfrom 7 to 12.3, particularly preferably from 7.5 to 10.5, veryparticularly preferably from 8.1 to 10.

The pH can be established by adding up to 3% by weight, based on theliquor, of lime (also calcium hydroxide). However, the amount-of limemay also be reduced to 0.3% by weight at the most.

In a preferred embodiment of the novel process, the use of lime isdispensed with. In the preferred embodiment, from 0.1 to 4% by weight ofone or more inorganic basic alkali metal compounds is added, for exampleone or more hydroxides or carbonates of alkali metals, preferably ofsodium or potassium, very particularly preferably of sodium. Othersuitable inorganic basic alkali metal compounds are alkali metalsilicates. Instead of basic alkali metal compounds, it is possible toadd magnesium oxide, magnesium hydroxide, amines, for example ammonia,methylamine, dimethylamine, ethylamine or triethylamine, or combinationsof alkali metal compound and one or more amines.

In addition to water, organic solvent may be present in the liquor, forexample up to 20% by volume of ethanol or isopropanol.

The novel process can be carried out in vessels in which liming isusually effected. Preferably, the novel process is carried out inrotatable drums having baffles. The speed is usually from 0.5 to100/min, preferably from 1.5 to 15/min, particularly preferably up to5/min. If liming is to be effected over a period of more than 8 hours,the speed is usually from 0.5 to 10/min, preferably 1.5 to 5/min,particularly preferably up to 3/min for 5 minutes within each hour, i.e.rotation for 5 minutes and a pause for 55 minutes per hour.

The pressure and temperature conditions for carrying out the novelprocess are generally not critical. It has proven suitable to carry itout at atmospheric pressure; a pressure up to 10 bar is alsoconceivable. Suitable temperatures are from 10 to 45° C., preferablyfrom 15 to 35° C., particularly preferably from 25 to 30° C.

The compound or the compounds of the formula I can be metered at thebeginning of the liming process, but the animal hides can first besoaked under basic conditions and one or more compounds of the formula Iand at least one compound which catalyzes the hydrolysis of peptidebonds can be metered in only after some time. The metering can beeffected in one step, i.e. the total amount of the compound or compoundsI used is metered in one step; compound I can also be metered inportions or continuously. It is also possible to employ at least onecompound which catalyzes the hydrolysis of peptide bonds. Compound I andthe compound which catalyzes the hydrolysis of peptide bonds can bemetered together or separately.

The novel process can be carried out within a period of from 5 minutesto 48 hours, preferably from 10 minutes to 36 hours, particularlypreferably from 20 minutes to 15 hours.

In one embodiment of the present invention, organic polyelectrolytes canbe added.

Organic polyelectrolytes are generally understood as meaning organicpolymers which have a large number of groups which are capable of ionicdissociation and may be an integral part of the polymer chains or may beattached as side groups to said chains. In general, each of the randomrepeating units carries at least one group capable of ionic dissociationin aqueous solution. In the present invention, ionomers are alsoincluded among the organic polyelectrolytes, said ionomers being thoseorganic polymers in which many repeating units carry a group capable ofionic dissociation, but not every unit carries such a group. Polymershaving only one or two ionizable groups at the respective chain ends or,in the case of branched polymers, a number of groups capable ofdissociation which corresponds to the number of chain ends are notincluded among polyelectrolytes in the context of the present invention.

In the novel process, polybases, polyacids, polyampholytes or thepolysalts or mixtures thereof may be used. Polyacids are to beunderstood as meaning those organic polyelectrolytes which dissociate inan aqueous medium with elimination of protons, for example havingpolyvinylsulfonic acid, polyvinylsulfuric acid, polyvinylphosphonicacid, polymethacrylic acid or polyacrylic acid as a random repeatingunit. Polybases are to be understood as meaning those organicpolyelectrolytes which contain groups or radicals which can beprotonated by reaction with Brönsted acids, for examplepolyethylenimines, polyvinylamines or polyvinylpyridines. Polyampholytesare usually understood as meaning those polymers which contain bothrepeating units which dissociate in an aqueous medium with eliminationof protons and repeating units which can be protonated by reaction withBrönsted acids. Polysalts are usually understood as meaning singly or inparticular multiply deprotonated polyacids.

Synthetic polyelectrolytes are preferably used in the novel process.

Of course, assistants customary in the tannery may also be added forcarrying out the novel process, for example phosphines, e.g.triphenylphosphine or tri(2-carboxyethyl)phosphine hydrochloride, andfurthermore hydroxylamine, urea, guanidine or guanidinium hydrochloride,hydrazine, biocides, surfactants and emulsifiers.

By means of the novel process, it is possible to produce excellentlyunhaired pelts. It is also found that the epidermis is completely or atleast substantially detached after only a short duration of treatment.It is furthermore found that, particularly in the treatment of animalhides of completely or partly black animals, for example Friesiancattle, a substantial proportion of melanin or even all melanin is alsodestroyed or removed from the pelts, so that particularly pale pelts areobtained. The present invention therefore relates to particularly palepelts, produced by the novel process.

It has furthermore been found that the pelts produced according to theinvention are very useful for the production of leather. After the novelpelts have been further processed by methods customary in the tannery,i.e. bating, if required deliming, pickling, chrome-free tanning orchrome tanning, retanning and finishing, it is observed that the peltsproduced according to the invention can be further processed to leatherwith an improved yield in terms of area and less swelling damagecompared with leather produced from pelts which were unhaired with theaid of, for example, Na₂S, NaHS, thioglycolic acid or aminoethanol.Moreover, it is possible to effect particularly level dyeing of thepelts produced according to the invention. If the use of lime isdispensed with in the novel process, novel pelts free of lime blast andhaving particularly flat and smooth grain are obtained.

Only slightly, preferably no, conversion to nubuck-type leather isobserved.

In a preferred embodiment, the bating step can be dispensed with in thefurther processing.

The present invention furthermore relates to leathers produced fromnovel pelts. Overall, they have advantageous performancecharacteristics.

It has furthermore been found that the wastewaters formed in the novelprocess, in particular wastewaters of novel processes in which Na₂S,NaSH or a mercaptan, such as aminoethanol or thioglycolic acid, is notemployed, can be particularly readily worked up. After the end of theaction of one or more compounds of the formula I and of one or morecompounds which catalyze the hydrolysis of peptide bonds on animalhides, the resulting pelts are separated from the liquor, for example bysimply taking out the pelts or by draining the liquor. The liquorseparated off is subsequently also referred to as novel residual liquoror as residual liquor. The residual liquor contains, inter alia, reactedand possibly unconsumed compound of the formula I, in addition to basicalkali metal compound or basic amines or lime and in particular residuesof the horny materials separated from the pelts and of the epidermis andmay contain melanin and/or degradation products of melanin. In apreferred embodiment, the novel residual liquor contains no noticeableamounts of compound of the formula I.

The present invention furthermore relates to residual liquors whichcontain only small amounts of Na₂S and preferably neither Na₂S nor NaHS,and, as organic sulfur compounds, those of the formula I and thereaction and secondary products thereof from the removal of hornysubstances from animal hides, and organic sulfur compounds whichoriginate from the animal hides. The novel residual liquors may nowcontain melanin and/or degradation products of melanin and melamineand/or degradation products of melamine. Moreover, the salt load isconsiderably reduced by using the process at a pH of less than 12.4, inparticular at a pH of from 7 to 10. This is possible in particular whenthe use of lime is dispensed with. The novel liquors are obtainable bythe novel process. In comparison with the residual tannery liquors knownfrom the prior art, they are virtually odorless and particularly simpleto work up.

The residual liquors contain reaction products and secondary products ofcompounds of the formula I which result from the removal of hornysubstances from the animal hides, mainly hydrolysis and oxidationproducts of compounds of the formula I, and proteins hydrolyzed with theaid of an organic compound.

It has been found that the novel residual liquors are particularly easyto work up.

The present invention furthermore relates to a process for working upnovel residual liquors. The novel working-up process comprises aplurality of steps.

In a first, optional step, the novel pelts are separated from the lime.This step is by its very nature required only when lime has been used inthe treatment of the animal hides, but is otherwise not required. Theseparation is effected by settling, flotation, decanting, filtration orcentrifuging, the separation of the lime by decanting, settling orfiltration being preferred in the case of large amounts of novelresidual liquors. By means of the first step described above, lime-freeresidual liquors are obtainable.

The lime-free residual liquors are then neutralized with acid, until apH of from 2 to 8, preferably from 3 to 7, particularly preferably from4 to 5, is reached.

Suitable acids are organic or inorganic acids. Examples are hydrochloricacid, phosphoric acid, CO₂, formic acid, sulfuric acid, acetic acid,citric acid, adipic acid and dicarboxylic acid mixtures comprisingadipic acid, glutaric acid and succinic acid. Acidification can beeffected without particular measures with regard to evolving hydrogensulfide.

The proteins removed during liming or during painting of the pelt areprecipitated or float, so that they are separated off mechanically in afurther step, for example by filtration or flotation.

It was furthermore found that, after neutralization and removal of theproteins, novel residual liquors can be used in an outstanding mannerfor soaking raw hides. The present invention therefore relates to theuse of the novel neutralized residual liquors freed from proteins as amedium for soaking raw hides. The present invention moreover relates toa process for working up novel residual liquors by neutralization andremoval of proteins.

The working examples which follow illustrate the invention.

GENERAL

Determination of the LVU

Hammarsten casein (commercially available from E. Merck, Art. 2242) inthe form of a 4% by weight solution was used.

A 4% by weight solution of casein was prepared by diluting 40 g ofcasein at up to 60° C. with 800 ml of distilled water and 32 ml of 1 NNaOH, mixing being effected until precipitates or undissolved solidswere no longer present. The solution was cooled to 25° C. and brought toa pH of 8.2 with 0.1 N NaOH or 0.1 N HCl. Dilution to 1 000 ml withdistilled water was then effected.

50 ml of the casein solution described above were mixed with 10 ml ofthe formulation to be investigated and comprising the compound(s) whichcatalyzes or catalyze the hydrolysis of peptide bonds, and the pH wasbrought to 8.2 with 0.1 N NaOH or 0.1 N HCl. After 60 minutes at 37° C.the reaction was stopped by adding 20 ml of 0.1 N HCl and 20 ml of 10%by weight Na₂SO₄ solution, any precipitate formed was filtered off and a20 ml sample was taken and titrated with 0.1 N NaOH againstphenolphthalein.

A blank sample was prepared by mixing the abovementioned reagentswithout adding the formulation to be investigated and comprising thecompound(s) which catalyzes or catalyze the hydrolysis of the peptidebonds. The further procedure was as described above. The differencebetween the consumptions of NaOH, multiplied by 17.39 and divided by theenzyme mass used in g, corresponds to the LVU/g.

Below, all data in % by weight are based on the salted weight, unlessstated otherwise.

General Working Methods 1. SOAKING

Soaking with the Use of Water

The salted South German cattle hide was first presoaked at 28° C. with150% by weight of water and 0.2% by weight of C₁₅H₃₁—O—(CH₂—CH₂—O)₇—Hfor 120 minutes in a drum with gentle agitation. The liquor wasdischarged (X1-1 Presoaking liquor, 200% by weight) and then-soaking waseffected with 100% by weight of water, 0.2% by weight ofC₁₅H₃₁—O—(CH₂—CH₂—O₇—H and 0.5% by weight of Na₂CO₃ with occasional agfor 19 hours. The liquor was then discharged (X1-2 Main soaking liquor,100% by weight).

2. HAIR-DESTROYING LIMING OF COMPARATIVE EXAMPLE V1

For comparative example V1, 100 parts by weight, based on salted weight,were treated in succession with 80 parts by weight of water and 1.0% byweight of mercaptoethanol in a rotatable 10 I drum containing baffles.After 30 minutes, 0.8% by weight of NaSH (70% by weight) and 1% byweight of calcium hydroxide followed for a further 30 minutes. 0.75% byweight of sodium sulfide and 0.75% by weight of sodium sulfide togetherwith 1.0% by weight of lime followed at an interval of 30 minutes. Thedrum was operated for a further 30 minutes at 15 revolutions/minute. Afurther 70 parts by weight and 1.0% by weight of lime were then metered.After 10 hours at from 23 to 27° C. and 5 minutes per hour at 3revolutions/minute, the experiments were terminated by discharging theliquor (sample V1-3 Liming liquor, 150% by weight) and washing the peltonce for 15 minutes with 150 parts by weight of water (sample V1-4Liming wash liquor, 150% by weight).

Before the further processing, the pelt was fleshed and split (2.8 mm).

2.1. Further Processing of the Pelt According to Comparative Example V1in the Deliming

Below, the data in % by weight were based on the pelt weight, grainsplit, 2.8 mm (corresponds to 75% of salted weight), unless statedotherwise. The deliming was carried out at from 25 to 32° C.Experimental parameters are shown in table 1.

TABLE 1 Experimental parameters of the further processing of the peltfrom VI Amount Experi- [% by Time ment wt.] Product pH [min] V1 150Water, 2× 20 Discharge liquor (V1-5/V1-6 Deliming wash liquor, 300% byweight) 100 Water 0.2 Deliming agent Decaltal ® ES-N, commerciallyavailable from BASF Aktiengesellschaft 0.2 C₁₅H₃₁—O—(CH₂CH₂—O)₇—H(diluted 1:3) 0.2 NaHSO₃ 8.6 20 Discharge liquor (V1-7 Deliming liquor,100% by weight) 50 Water 1.0 Deliming agent Decaltal ® ES-N, 8.0 45commercially available from BASF Aktiengesellschaft 1.0 Basozym ® C10,1000 LVU/g 45 Discharge liquor (V1-8 Bating liquor, 50% by weight) 150Water 10 Discharge liquor (V1-9 Bating wash liquor, 100% by weight)

The penetration of the neutralization over the hide cross section waschecked using phenolphthalein as indicator. The time required for thispurpose was noted.

2.2. Pickling and Tanning of the Pelt According to Comparative ExampleV1

Below, the data in % by weight are based on the pelt weight, grainsplit, 2.8 mm (corresponds to 75% of salted weight) unless statedotherwise.

40% by weight of water and 6% by weight of NaCl (8° Be) were added to100% by weight of the respective novel pelt E1 to E6 in a rotatable 10 Idrum containing baffles. After 10 minutes, 1.0% by weight of thefatliquoring agent Lipoderm Licker® A1, commercially available from BASFAktiengesellschaft, was added and, after 20 minutes, 0.4% by weight ofaqueous formic acid (20% by weight) was introduced. After 30 minutes,0.8% by weight of 98% by weight sulfuric acid was added; the pH was 3.0.After a further 90 minutes, 2.5% by weight of a dispersion of Relugan®GTP, diluted in the volume ratio of 1:3 with water, 3.0% by weight of adispersion of the syntan tanning agent Basyntan® SW liquid diluted inthe volume ratio 1:2 with water (both reagents commercially availablefrom BASF Aktiengesellschaft) and 2.0% by weight of anaphthalenesulfonic acid/formaldehyde condensate, prepared according toU.S. Pat. No. 5,186,846, example Dispersant 1, were added. Saidsubstances were allowed to act for 90 minutes with occasional rotationand basification was effected with 0.2% by weight of sodium formate to apH of 3.9. After a contact time of 15 hours, a further 0.2% by weight ofsodium formate and 0.2% by weight of NaHCO₃ were added. The pH was now4.0. After a further 90 minutes, 0.2% by weight of a dispersion of thefungicide Cortymol® Fun, diluted in the volume ratio of 1:3 with water,was added.

After the end of the action, the residual liquor was discharged, and theresidual liquor V-0 was obtained.

3. EXAMPLES E1 to E7 ACCORDING TO THE INVENTION 3.1. Hair-destroyingLiming of Examples E1 to E7 According to the Invention

For examples E1 to E5 according to the invention, 100 parts by weight,based on salted weight, were treated with 50 parts by weight of water ina rotatable 10 I drum containing baffles. The drum was rotated.Thereafter, the amount of enzyme shown in table 2 and, after 60 minutes,the amount of racemic dithiothreitol (DTT) shown in table 2 were added.The pH was 7.5. After the time shown in table 2, NaOH solution (50% byweight in water) was added as Base 1. The pH increased to the valuedstated in the table. In examples E1 to E5, in each case the amount ofNaOH solution (50% by weight in water) stated in table 2 was alsometered in as Base 2, the pH increasing to the value stated in thetable.

The drum was rotated in each case for 5 minutes at 5 rpm and leftstationary for 55 minutes, after which the movement cycle was repeated.After a contact time of 10 hours, 50% by weight of water were added, theliquor was discharged, 150% by weight of water were introduced, movementwas effected for 10 minutes and the wash liquor was again discharged.

Example E6 was carried out analogously, except that NaOH solution wasreplaced by solid MgO and the subsequent metering of base was dispensedwith.

Example E7 was carried out analogously, except that the addition of basewas dispensed with.

TABLE 2 Experimental parameters of liming of the examples according tothe invention Example E1 E2 E3 E4 E5 E6 E7 Enzyme Basozym PyraseAlcalase Alcalase Basozym Basozym Basozym L10 250 mp 3.0t 3.0t L10 L10L10 (a) and Basozym S20 (b) Amount of 2.0 0.006 0.016 0.008 1.0 2.0  1.5(a) enzyme  0.4 (b) [% by wt.] LVU/g 1000 350000 250000 250000 1000 1000 1000 (a)  2000 (b) DTT 0.75 0.75 0.75 0.75 1.5 0.75 1.5 [% by wt.] Base1 1.5 1.5 1.5 2.0 2.0 1.0 — [% by wt.] pH 10.7 10.7 10.7 12.1 12.1 10.58.5 Time[h] 3 3 3 0.5 0.5 — 0.5 Base 2 1.5 1.5 1.5 0.4 0.4 — — [% bywt.] pH 12.4 12.4 12.4 12.4 12.4 10.5 8.5 Liming E1-3 E2-3 E3-3 E4-3E5-3 E6-3 E7-3 liquor Liming E1-4 E2-4 E3-4 E4-4 E5-4 E6-4 E7-4 washliquor

Basozym L 10 occasionally referred to as Basyzym L 10; proteolyticenzyme preparation containing 1000 LVU/g.

The novel pelts B E1 to B E7 were obtained.

Before the further processing, the pelts were fleshed and split (2.8mm).

3.2. Pickling and Tanning of the Pelts of Examples E1 to E7 According tothe Invention

Below, the data in % by weight are based on the pelt weight, grainsplit, 2.8 mm (corresponds to 75% of salted weight), unless statedotherwise.

40% by weight of water and 6% by weight of NaCl (8° Be) were added to100% by weight of the respective novel pelt E1 to E6 in a rotatable 10 Idrum containing baffles. After 10 minutes, 1.0% by weight of thefatliquoring agent Lipoderm Licker® A1, commercially available from BASFAktiengesellschaft, was added and, after 20 minutes, 0.4% by weight ofaqueous formic acid schaft and, after 20 minutes 0.4% by weight ofaqueous formic acid (20% by weight) was introduced. After 30 minutes,0.8% by weight of 98% by weight sulfuric acid was added; the pH was 3.0.After a further 90 minutes, 2.5% by weight of a dispersion of theleather dye Relugan® GTP, diluted in the volume ratio of 1:3 with water,3.0% by weight of a dispersion of the syntan tanning agent Basyntan® SWliquid diluted in the volume ratio 1:2 with water (both reagentscommercially available from BASF Aktiengesellschaft) and 2.0% by weightof a naphthalenesulfonic acid/formaldehyde condensate, preparedaccording to U.S. Pat. No. 5,186,846, example Dispersant 1, were added.Said substances were allowed to act for 90 minutes with occasionalrotation and basification was effected with 0.2% by weight of sodiumformate to a pH of 3.9. After a contact time of 15 hours, a further 0.2%by weight of sodium formate and 0.2% by weight of NaHCO₃ were added. ThepH was now 4.0. After a further 90 minutes, 0.2% by weight of adispersion of the fungicide Cortymol® Fun, diluted in the volume ratioof 1:3 with water, was added.

After the end of the action, the residual liquor was discharged, and theresidual liquors E1-5 to E6-5 and the novel leathers L E1 to L E6 wereobtained.

4. ASSESSMENT OF THE PELTS ACCORDING TO COMPARATIVE EXAMPLE B V1 AND OFEXAMPLES B E1 TO B E7 ACCORDING TO THE INVENTION AND OF THE LEATHERSACCORDING TO COMPARATIVE EXAMPLES L V1 AND ACCORDING TO EXAMPLES L E1 TOL E7 ACCORDING TO THE INVENTION

The leathers produced according to the invention have a smoother andflatter grain, without visible conversion to nubuck-type leather, incomparison with the leather according to the comparative example.

The epidermis and the hairs with hair root had been completely removedfrom the pelts of the examples according to the invention or destroyed.Particularly striking and advantageous was the very pale appearance ofthe novel pelts. The bluish shadows customary for lime/sodium sulfideliming (reaction of sulfide with iron ions) and lime blast, which canlead to nonlevel dyeings, in particular in the case of pale hues, werecompletely absent. Furthermore, the properties of the pelts producedaccording to the invention were excellent with respect to swelling.

5. FURTHER PROCESSING OF THE LEATHER ACCORDING TO COMPARATIVE EXAMPLE LV1 AND ACCORDING TO EXAMPLES L E1 TO L E7 ACCORDING TO THE INVENTION INRETANNING

The following polymers were used:

Polymer 1: Alternating copolymer of (C₂₀-C₂₄-α-olefin)/maleic anhydride;molar comonomer fraction of the (sum of the α-olefins): maleic anhydride1:1, M_(w) 8 900 g, preparation described in EP 0 412 389 B1 asdispersion I. Form used: 30.2% by weight of dispersion.

Polymer 2: 30% strength by weight aqueous polymer solution partlyneutralized with NaOH; homopolymer of methacrylic acid, M_(n) about 10000 g/mol; Fikentscher K value: 12, viscosity of the 30% by weightsolution: 65 mPa.s (DIN EN ISO 3219, 23° C.), pH 5.1.

The leathers obtained according to 3. were sammed and shaved byconventional methods. The shaved thickness of the leathers was 2.0-2.2mm (shaved weight corresponds to 25% of salted weight). The retanningwas effected as follows:

The pretanned leather L V1 or L E1 to L E6 was treated, at a liquorlength of 100% by weight of water at 30° C., with 15% by weight ofpolymer 1 as a 30.2% by weight aqueous dispersion and 15% by weight of a30% by weight aqueous dispersion of polymer 2 (action step (a), cf.table 4). Thereafter, the commercial dye Luganil® Black AS liquid wasadded to the leather. A further 10% by weight of polymer 1 in the formof 30.2% strength by weight aqueous dispersion and 2% by weight ofpolymer 2 in the form of a 20% by weight dispersion were then added. Theleather remained for the time stated in table 4 in the liquor thusformed (action step (b)).

The reaction temperature was then increased by adding 100% of water at45° C. A pH of 3.5 was established with formic acid. The leatherremained for the time stated in table 4 in the liquor thus formed(action step (c).

Finally, the leather was dyed with a solution of 1.5% by weight ofLuganil® Black AS liquid in 100% by weight of water and 0.7% by weightof formic acid over a period of 45 minutes, and then washed, fixed andfinished in the customary manner. The finished crust leathers C V1(comparative example) and C E1 to C E6 (according to the invention) wereobtained.

The process parameters are shown in table 3.

TABLE 3 Process parameters of the action steps in the retanning L V1, LE1 to L E7 Action step (a) Polymers 1 and 2 Duration  90 min Action step(b) Polymers 1 and 2 Duration 180 min Action step (c) Polymer — Duration 20 min

The physical properties and the performance characteristics were thentested.

6. ASSESSMENT OF THE FINISHED LEATHERS C V1 AND C E1 TO C E6

The crust leathers produced from the examples according to the inventiondiffer from the comparative example in their haptic and opticalproperties through the smoother and finer grains. Leather having verygood dyeing and tight grain in combination with very good body andexcellent softness with elegant handle is obtained. The values are shownin table 4.

TABLE 4 Performance characteristics of crust leathers C V1 and C E1 to CE6 Stitch tear resistance according to Unhairing Grain tightness DIN53331 Example activity Wet white leather [N] C V1 2 2 140 C El 1 1 172 CE2 1 1 178 C E3 1 1 180 C E4 1 1 187 C E5 1 1 190 C E6 1 1 181 C E7 1 1195

The unhairing activity and the grain tightness were assessed visuallyusing ratings from 1 (very good) to 6 (inadequate).

7. WORKING UP THE RESIDUAL LIQUORS

General working method using the example of the residual liquorsaccording to example E1

The liming liquor E1-3 and liming wash liquor E1-4 were combined andwere brought to a pH of 4.5 with concentrated sulfuric acid (98% byweight). The precipitated protein was separated off using a chamberfilter press. The data of the combined and purified liquors E1-3 andE1-4 are shown under 8.1 (liquor E1-A). The purified liming liquors werevery useful as soaking liquors. The water consumption can beconsiderably reduced therewith.

The residual liquors of the examples according to the invention could beacidified to a pH of 4.5 with sulfuric acid without evolution ofhydrogen sulfide, and the precipitated protein could be separated offwithout problems by filtration. The residual liquors were moreovervirtually clear.

The liquor according to comparative experiment V1 could not be acidifiedwithout precautions and evolved foul-smelling hydrogen sulfide. Evenafter working up, it could not be used for soaking cattle hides.

8. ANALYTICAL RESULTS OF THE RESIDUAL LIQUORS AND WASTEWATERS

TABLE 5 Analytical results of the residual liquors and wastewater Waterconsumption Water consumption COD COD_(total) COD Experiment up totanning [m³] up to tanning [rel. %] [mg O₂/l] [kg O₂] [rel. %] V1 10.30100 13200 136.2 100 E1 2.80 27 15500 43.4 31 E2 2.80 27 16800 47.0 34 E32.80 27 18900 52.9 38 E4 2.80 27 18900 52.9 38 E5 2.80 27 19000 53.2 38E6 2.80 27 17300 48.4 35 E7 2.80 27 18700 52.8 38 COD: chemical oxygendemand

8.1. Protein Precipitate According to E1-E6

In each case about 100-150 kg, solids content 30% by weight, COD [kgO₂/kg]83.3-92.8, ash content 1.0-1.4%

8.2. Worked-Up and Reused Liquor Liquors for Example E1

TABLE 6 Analytical values of the worked-up residual liquors E1-A Solidscon- Ash Liquor tent [% by COD [% by Liquor Process pH [%] wt.] [mgO₂/l] wt.] E1-A E1-3 + E1-4 12.4 7.8 0.8 44300 250 (beforeneutralization) E1-A E1-3 + E1-4 4.5 5.7 2.6 6200 250 (afterneutralization, after filtration)

8.3. Use of Neutralized Residual Liquors Freed from Protein

Soaking Using Neutralized Residual Liquors Freed from Protein

Method 1.1 was repeated with a salted South German cattle hide, butwater was replaced by the neutralized residual liquor freed from proteinand described under 7.

The soaked hide was then further processed similarly to E1. A crustleather having the same properties as C E1 was obtained.

1. A process for removing horny substances from an animal hide,comprising: treating an animal hide with an aqueous liquor, wherebyhorny substances are removed from said animal hide, and wherein saidaqueous liquor has a pH of from 7 to 12.3 and contains lime which ispresent in an amount not greater than 0.3% by weight, and which aqueousliquor comprises from 0.05 to 5% by weight, based on the salted weightof said animal hide, of one or more compounds of the following formulaI:

or a corresponding alkali metal or alkaline earth metal or ammonium orphosphonium salts thereof, the variables being defined as follows: R¹ isselected from the group consisting of hydrogen and C₁-C₁₂-alkyl,unsubstituted or substituted by one or more S—H or O—H groups; X¹ to X⁴are identical or different and are selected from the group consisting ofhydrogen, C₁-C₄-alkyl, O—H, S—H and N—R²; R² is hydrogen or C₁-C₁₂-alkylor a C₁-C₄-alkyl-C═O group; at least one of the radicals X¹ to X⁴ beingS—H; and, if R¹ contains neither O—H nor S—H, at least one furtherradical from among X¹ to X⁴ is selected from the group consisting ofS—H, OH and NH−R²; and further comprising at least one compound whichcatalyzes a hydrolysis of peptide bonds.
 2. The process as claimed inclaim 1, wherein said at least one compound which catalyzes saidhydrolysis of peptide bonds is an organic compound.
 3. The process asclaimed in claim 1, wherein said at least one compound which catalyzessaid hydrolysis of peptide bonds is an enzyme.
 4. The process as claimedin claim 3, wherein said enzyme is a protease or a peptidase.
 5. Theprocess as claimed in claim 1, wherein said at least one compound whichcatalyzes said hydrolysis of peptide bonds has Löhlein and Volhardactivity from 500 to 2,000,000 LVU/kg, based on the salted weight ofsaid animal hide.
 6. The process as claimed in claim 1, wherein R¹ is H,X¹ and X⁴ are S—H, and X² and X³ are O—H.
 7. The process as claimed inclaim 1, wherein the amount of said at least one compound whichcatalyzes said hydrolysis of peptide bonds is chosen to be a factor ofat least 10 smaller than the amount of said one or more compounds offormula I.
 8. The process as claimed in claim 1, wherein said process iscarried out in the presence of urea.
 9. The process as claimed in claim1, further comprising the steps of obtaining a residual liquor, whereinthe residual liquor is separated off from lime and then neutralized withacid and then, proteins are separated off.
 10. A process for soaking rawhides comprising soaking a raw hide with said neutralized residualliquor as claimed in claim
 9. 11. The process as claimed in claim 1,wherein the pH is from 7.5 to 10.5.
 12. The process as claimed in claim1, wherein the pH is from 8.1 to
 10. 13. The process as claimed in claim1, wherein said animal hide contains an epidermis, and at least partialdetachment of the epidermis from said animal hide is carried out by saidprocess.
 14. A process for removing horny substances from an animalhide, comprising: treating an animal hide with an aqueous liquor,whereby horny substances are removed from said animal hide, and whereinsaid aqueous liquor contains lime which is present in an amount notgreater than 0.3% by weight, and which aqueous liquor comprises from0.05 to 5% by weight, based on the salted weight of said animal hide, ofone or more compounds of the following formula I:

or a corresponding alkali metal or alkaline earth metal or ammonium orphosphonium salts thereof, the variables being defined as follows: R¹ isselected from the group consisting of hydrogen and C₁-C₁₂-alkyl,unsubstituted or substituted by one or more S—H or O—H groups; X¹ to X⁴are identical or different and are selected from the group consisting ofhydrogen, C₁-C₄-alkyl, O—H, S—H and N—HR²; R² is hydrogen orC₁-C₁₂-alkyl or a C₁-C₄-alkyl-C═O group; at least one of the radicals X¹to X⁴ being S—H; and, if R¹ contains neither O—H nor S—H, at least onefurther radical from among X¹ to X⁴ is selected from the groupconsisting of S—H, OH and NH—R²; and further comprising at least onecompound which catalyzes a hydrolysis of peptide bonds, wherein theamount of said at least one compound which catalyzes said hydrolysis ofpeptide bonds is chosen to be a factor of at least 10 smaller than theamount of said one or more compounds of formula I.